One of the goals of synthetic biology is to be able to engineer cells that can sense and report on new signals. Cells that could be modified to sense specified cell-cell contacts would be extremely useful for a wide range of biomedical and research applications, ranging from mapping of cell connectivity networks to controlling the exact location of stem cell differentiation for tissue repair or regeneration. In preliminary studis, we have recently developed a new synthetic receptor construct, based on the Notch receptor, in which we can swap the extracellular ligands that are sensed by the receptor. This receptor, when activated by contact with a cell expressing the cognate ligand (but not soluble ligand), induces a user-defined transcriptional response. Thus this engineered receptor platform can in principle be used to flexibly link a wide variety of cell-contact stimuli to a flexible, modular response. I plan to characterize and show two applications of this receptor platform. During the K99 phase I will carry out two aims. One is to optimize and mechanistically characterize the synthetic cell-cell contact receptor system, by characterizing its modularity. For this aim, I will use protein- engineering techniques to build libraries of receptor variants; I will characterize th receptor by expressing these libraries in fibroblasts cells and monitor reporter transcription by fluorescent activation cell sorter (FACS). The second aim is to use this synthetic receptor platform as a toolkit to map cell-cell connectivity. For this aim, I will use the libraries of the irst aim and follow the contact-dependent activation of reporter genes by live microscopy. I plan also to extend the combinatorial power of the system by implementing logic gate and temporal control over receptor response. For the R00 phase, I plan to develop a third aim: use this synthetic receptor platform as a way to control cell differentiation in response to nucleating cell or surface scaffold. I will engineer mouse embryonic stem cells with chimeric receptors to differentiate only when contacted by ligands presented by neighbors or by scaffold. These studies will represent a foundation for the design of a new wave of powerful tools for the control of cell behavior via cell-cell and cell-surface contacts. The possibility of reporting and controllng cell behavior through these receptors will establish new paradigms for basic research, cell therapy and tissue engineering. The successful characterization of this receptor platform should give us a customizable and highly flexible toolkit to control cell behavior, as well as to map complex relationships between cells in live tissue. In the future we will be able to report on complex cell-cell contacts and distinguish populations of cells based on their neighbor relationships in vivo, in neurons and other developmental contexts. These tools will also enable the control of complex differentiation programs of ES cells both in vitro to increase our fundamental understanding, and in vivo to control the exact location of stem cell differentiation for tissue repair or regeneration, perhaps responding specifically to signals associated with tissue damage or injury.